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SUNSCREEN AND ANTI-OXIDANT ACTIVITY OF HERBAL GEL OF
CUCUMBER EXTRACT
Garima Garg*1, Shivani Singh
2 and Vipin Kumar Garg
1
1Department of Pharmaceutical Technology, Meerut Institute of Engineering & Technology,
NH-58, Baghpat By-Pass Crossing, Delhi-Haridwar Highway, Meerut-250005, India.
2Llyod College of Pharmaceutical Science and Research, Greater Noida. India.
ABSTRACT
As in daily life humans are exposed to harmful UV radiations which
cause tanning, sunburns, edema and various other allergic reactions it
becomes a need to use such agents which can protect the human body
from the above mentioned side effects. Since it is more beneficial to
use natural agents in comparison to synthetic ones we have taken
cucumber as an ingredient to prepare a sunscreen gel and evaluated it
for its effectiveness as a sunscreen preparation. The formulated gel was
evaluated for its SPF (Sun Protection Factor) i.e. protection against
UVB rays and UVA: UVB ratio i.e. protection against UVA rays and it
was found that the cucumber gel has potential to provide protection
against both UVA rays and UVB rays. Additionally it was also found
to have antioxidant properties which may contribute to sunscreen
activity. The gel was found to have good spreadability, homogeneity
and pH within the range of skin pH and thus suitable to use on skin.
KEYWORDS: Sun Protection Factor (SPF), Boot star rating, DPPH, UVA/UVB ratio
INTRODUCTION
Cucumber, scientifically known as Cucumis sativus (family- cucurbitaceae) are obtained
throughout India. The flesh of cucumber is primarily composed of water but also contains
ascorbic acid (vitamin C) and caffeic acid, both of which help soothe skin irritations and
reduce swelling. The silica in cucumber is an essential component of healthy connective
tissue, which includes muscles, tendons, ligaments, cartilage and bone.[1]
Cucumber juice is a
source of silica which can improve the skin complexion and health and also give natural
World Journal of Pharmaceutical Research SJIF Impact Factor 6.805
Volume 5, Issue 7, 1202-1213. Research Article ISSN 2277– 7105
*Corresponding Author
Garima Garg
Department of
Pharmaceutical Technology,
Meerut Institute of
Engineering & Technology,
NH-58, Baghpat By-Pass
Crossing, Delhi-Haridwar
Highway, Meerut-250005,
India.
Article Received on
15 May 2016,
Revised on 05 June 2016,
Accepted on 26 June 2016
DOI: 10.20959/wjpr20167-6542
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Garg et al. World Journal of Pharmaceutical Research
glow to skin due to its hydrating property. Cucumber had been used traditionally for swelling
under the eyes, dark circles and sunburn etc due to two compounds, ascorbic acid and caffeic
acid which prevent water retention. The present work deals with the preparation and
evaluation of cucumber gel for antioxidant and sunscreen activity.
MATERIALS AND METHODS
Plant Material
Fresh cucumbers were obtained from the local market of Meerut. DPPH was obtained from
Sigma Aldrich chemical ltd Germany. Carbopol 940 LR was purchased from SD Fine
Chemical Ltd. Mumbai. Benzophenone, ferric chloride and trichloroacetic acid were
purchased from Central Drug House, New Delhi. Potassium ferricyanide was purchased from
RFCL Ltd New Delhi. All other chemicals used were of analytical grade.
PREPARATION OF EXTRACT
For preparation of cucumber extract, cucumbers were properly peeled, washed and chopped
finely. The chopped material was then crushed in a grinder and the slurry obtained was
passed through the muslin cloth in order to obtain the cucumber juice. The juice was then
lyophilized at a temperature of -700C so that a freeze dried extract in the solid form can be
obtained. The extracts were stored by keeping in refrigerator till further use.
PREPARATION OF GEL
For preparing the gel carbopol- 940 LR was soaked in water for 5-6 hours to obtain a
homogenous mass. The extract was then incorporated in carbopol and triturated well. All the
other ingredients were added and mixed well.[2]
TABLE I: Formulation of Gel
S.No. Ingredient Quantity
1 Carbopol 1
2 Herbal extract 2.5gm
3 Triethanolamine To pH 6-7
4 Glycerine 5%
5 Distilled water Upto 25 ml
EVALUATION OF SUNSCREEN ACTIVITY
1. Benzophenone method[3]
For the measurement of SPF (Sun Protection Factor) the effectiveness of sunscreen gels were
analysed using the reaction to produce benzopinacol. Underlying principle is that reaction of
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benzophenone with isopropanol depends on the absorption of UV light. The conversion
products include benzopinacol and acetone. The benzopinacol is the insoluble product that is
removed when the solution is filtered. The more UV light that reaches the solution of
benzophenone and isopropyl alcohol, the greater the amount of benzopinacol precipitate
formed. Therefore, the difference in yield between a vial coated with sunscreen gel and one
without will be proportional to the reduction in absorbed UV radiation caused by the
sunscreen gel.
+ CH
OH
H3C CH3
uv radiation
Isopropanol
CHO OH+
C
O
CH3 CH3
Benzopinacol
Acetone
C
Fig 1: Mechanism of Benzophenone Method for determination of SPF
Benzophenone solution
20% w/v solution of benzophenone was prepared by dissolving 4gms of benzophenone in 20
ml of isopropanol.
Study Design
For the analysis of sunscreen activity of the laboratory prepared gel the product was
compared to the commercial preparations of varying SPF 15, 20, 30 and a control in which no
preparation is applied. Five Petri plates were taken and 10ml of solution was added to each of
them. After adding the solvent the petri plates were covered properly and uniform layer of
different preparations were applied. One petri plate which does not contain any preparation
was considered as control. The petri plates were kept in sunlight for a period of 6 hours and
thereafter kept in dark for overnight and readings were recorded as the amount of crystals of
benzopinacol.
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2. UV spectrophotometric method[4]
The in vitro methods are in general of two types. a) Methods which involve the measurement
of absorption or the transmission of UV radiation through sunscreen product films in quartz
plates or biomembranes. b) Methods in which the absorption characteristics of the sunscreens
agents are determined based on spectrophotometric analysis of dilute solutions. Mansur et al.
(1986) developed a very simple mathematical equation utilizing UV spectrophotometry. The
following equation was given by Mansur
SPFspectrophotometric = CF x320
∑290 EE (λ) x I(λ) x Abs(λ) ……………….(1)
Where EE – erythemal effect spectrum, I – solar intensity spectrum, Abs - absorbance of
sunscreen product; CF – correction factor (= 10). The values of EE x I are constants. They
were determined by Sayre et al. (1979) and are showed in Table II. The aim of this research
was to determine the SPF values of sunscreens gel containing natural plant extracts by UV
spectrophotometry applying Mansur mathematical equation (Equation 1).
TABLE II: Normalized product function used in the calculation of SPF (Sayre et al.,
1979)
Wavelength(λ)nm EE X I (normalized)
290 0.0150
295 0.0817
300 0.2874
305 0.3278
310 0.1864
315 0.0839
320 0.0180
Total 1
Method
1.0 g of all samples was weighed, transferred to a 100 ml volumetric flask, diluted to volume
with water, followed by ultrasonication for 5 min and then filtered through cotton, rejecting
the ten first ml. A 5.0 ml aliquot was transferred to 50 ml volumetric flask and diluted to
volume with water.
The absorption spectra of samples in solution were obtained in the range of 290 to 450 nm
using 1 cm quartz cell and water as a blank. The absorption data were obtained in the range
of 290 to 320, every 5 nm and 3 determinations were made at each point, followed by the
application of Mansur equation.
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3. BOOT STAR rating system[5]
The UVA: UVB ratio used in this UVA labelling system is calculated as an indicator of the
UVA absorbance properties of a sunscreen product relative to UVB, rather than a ratio of
protection factors. It gives an indication of the physical properties of the sunscreen. UVA:
UVB ratios determined using absorbance measurements rather than MPF values will be less
affected by either "in use" application rate or variations in application rate during the in-vitro
UVA evaluation technique. This is because absorbance values are directly proportional to
film thickness and concentration of active ingredient whereas protection factors are not. (An
increase in product application rate does not necessarily result in the same factor increase in
SPF value.
The index of UVA protection is calculated from a plot of wavelength versus log MPF (ie
absorbance). It is defined as the area (per unit wavelength) under the UVA portion of the plot,
divided by the area (per unit wavelength) under the UVB portion of the same curve.
UVA area per unit λ =
UVA area per unit λ =
The integrals in the above formulae can be approximated by using Simpson's Rule for
irregular areas which states:-
Area = 1/3h x [(Y0 + Y2m) + 4(Y1 + Y3 + Y5 … + Y2m-1) + 2(Y2 + Y4 + Y6 … + Y2m-2)].
Where Y0, Y1, Y2, Y3…..Y2m are the lengths of a number, ‘2m’ of parallel lines drawn
vertically to divide the area under the curve of a graph into ‘2m – 1’ segments of equal width,
‘h’. In practice, the values of Y0, Y1, Y2, Y3…..Y2m are the absorbance values (aλ) measured
for the sunscreen product and ‘h’ is the wavelength interval at which the absorbance values or
MPFs are determined - eg 5 nm.
UVA area per unit wavelength (aUVA/λ)
aUVA/λ = 1/3 x 5 x [(a320 + a400) + 4 (a325 + a330 + a335 ….. + a395) + 2 (a330 + a340 + a350 ….. +
a390)] / 80.
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UVB area per unit wavelength (aUVB/λ)
aUVB/λ= 1/3 x 5 x [(a290 + a320) + 4 (a295 + a305 + a315) + 2 (a300 + a 310)] / 30.
The UVA: UVB ratio is then calculated as:-
The calculation of the UVA: UVB absorbance ratio will typically yield values from zero
(equal to no UVA absorbance) up to 1.0 (UVA absorbance equal to UVB). Once a UVA:
UVB ratio has been determined for a sunscreen product according to the above procedures,
the sunscreen can be categorised according to the Boots UVA Star Rating System below.
TABLE III: BOOT STAR rating system
Mean UVA: UVB Ratio
Star Rating
Category
Category
Descriptor
2nm or greater than
2nm increment
measurement
Less than 2nm increment
measurement
0.0 to 0.19 0.0 to 0.2 - No claim
0.2 to 0.39 0.21 to 0.4 * Minimum
0.4 to 0.59 0.41 to 0.6 ** Moderate
0.6 to 0.79 0.61 to 0.8 *** Good
0.8 to 0.89 0.81 to 0.9 **** Superior
0.9and above 0.91 and above ***** Ultra
MEASUREMENT OF ANTIOXIDANT ACTIVITY
1. Determination of reducing power[6,7]
The antioxidant activity of cucumber extract was determined using UV spectrophotometric
method and ascorbic acid as a standard. Extract and ascorbic acid were dissolved separately
in 1ml of double distilled water in a volumetric flask. To each of the volumetric flask 2.5 ml
of 0.2M phosphate buffer (pH 6.6) and 2.5ml of potassium ferricyanide was added.
The mixture was incubated in oven for 20 min at 500c. To the incubated solutions 2.5ml of
10% trichloroacetic acid was added and centrifuged for 10 min at 3000 rpm. From the above
solution 2.5ml of solution was taken out without disturbing and to it 2.5ml of distilled water
and 0.5ml of 1% freshly prepared Fecl3 was added. Absorbance was recorded at 700nm.
2. Determination of DPPH radical scavenging activity[8]
The free radical scavenging activity of the Cucumis sativus extract and ascorbic acid was
measured. 0.1 mM solution of DPPH in ethanol was prepared and 1.0 ml of this solution was
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added to 3.0 ml of aqueous solution of extract at different concentrations (1-9 mg/ml). Thirty
minutes later, the absorbance was measured at 517 nm. Lower absorbance of the reaction
mixture indicates higher free radical scavenging activity. The capability to scavenge the
DPPH radical was calculated using the following equation:
Where Acont = absorbance of the control reaction and Atest = absorbance in the presence of
the sample of the extract. The antioxidant activity of the extract was expressed as IC50. The
IC50 value was defined as the concentration (in μg/ml) of extracts that inhibits the formation
of DPPH radicals by 50%.
pH[9]
The pH of the prepared gel formulation was determined by using digital pH meter.
Spreadability[9]
Spreadability was observed by spreading 1 g of formulation on a clean even glass surface.
Homogeneity[10]
The prepared gel was tested for homogeneity by visual inspection after the gels have been set
in the container. They were tested for their appearance and presence of any aggregates.
Swelling Index Study of Topical Gel[10]
Swelling of the polymer depends on the concentration of the polymer, ionic strength and the
presence of water. To determine the swelling index of prepared topical gel, 1 gm of gel was
taken on porous aluminum foil and then placed separately in a 50 ml beaker containing 10 ml
water. Then samples were removed from beakers at different time intervals and put it on dry
place for some time after it reweighed. Swelling index was calculated as follows:
Swelling Index (SW) % = [(Wt – Wo) / Wo] × 100.
Wt = final weight of swollen gel at time t, Wo = initial weight of gel.
RESULTS AND DISCUSSION
Sunscreen Activity
1. Benzophenone method
This method compares the ability of different products to block UV radiations. Less the
amount of UV reaching to the benzophenone-isopropanol solution less will be the amount of
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crystal formed and more will be the sunscreen activity of product. The ability of the prepared
cucumber gel was compared with that of various marketed sunscreen preparations having
different SPF. The results obtained are shown in the Table IV. From the observations the SPF
of cucumber gel was found to be 3.191465.
TABLE IV: Determination of SPF by benzophenone method
S.No. Formulation Yield (in gm) SPF
1 Control 1.39 0
2 SPF 15 0.79 15
3 SPF 20 0.64 20
4 SPF 30 0.39 30
5 Cucumber gel 1.25 3.191465
2. UV spectrophotometric method
The value of sun protection factor of the cucumber gel was also calculated by the UV
spectrophotometric method. The absorbance of the sample was determined in range from
290-400nm at 5nm wavelength interval. The SPF is mainly the measure of protection against
UVB rays having range between 290-320 nm so we calculated the SPF value over this range.
The SPF of the formulation by this method was found to be 2.95.
TABLE V: Determination of SPF by UV spectrophotometric method
S.No. Wavelength(λ)nm EE X I
(normalized) Absorbance SPF
1 290 0.0150 0.015 0.05175
2 295 0.0817 0.0817 0.265525
3 300 0.2874 0.2874 0.885192
4 305 0.3278 0.3278 0.957176
5 310 0.1864 0.1864 0.520056
6 315 0.0839 0.0839 0.222335
7 320 0.0180 0.018 0.04572
Total 1 2.947754
3. BOOT STAR RATING System
As the boot star rating system specifies the category of the formulation with reference to the
protection against UVA rays in comparison to UVB rays, the value of UVA/UVB ratio
determines the protection against UVA rays and it was observed that the formulation has the
UVA/UVB ratio value of 0.666 [Table VI] and thus have a 3 star rating and good protection.
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TABLE VI: BOOT STAR rating for cucumber gel
S.No.
UVA area
per unit
wave length
UVB area
per unit
wave length
UVA:UVB
ratio
BOOT
STAR
rating
Category
descriptor
1 0.196 0.294 0.666 *** Good
ANTIOXIDANT ACTIVITY
1. Determination of reducing power
The antioxidant activity of cucumber gel was measured in terms of reducing power using
ascorbic acid as a standard. Antioxidant activity of ascorbic acid and cucumber gel was found
to be 100% and 30.83% respectively [Table VII].
TABLE VII: Determination of reducing power
S.No. Sample Absorbance
(700nm) Antioxidant activity
1 Ascorbic acid 2.4 100%
2 Cucumber extract 0.74 30.83%
2. Determination of DPPH radical scavenging activity
The gel formulation showed concentration dependent anti-radical activity by inhibiting DPPH
radical. Table VIII exhibits % DPPH scavenging activity and IC50 value (concentration in
μg/ml of gel formulations that inhibits the formation of DPPH radical by 50%) of the gel
formulation was found to be 6.44. Figure 1 shows the %DPPH scavenging activity.
TABLE VIII: Determination of DPPH radical scavenging activity.
S.No. Concentration
(mg/ml)
% DPPH scavenging activity
Cucumber gel Ascorbic acid
1 1 15.17094 33.54701
2 2 19.65812 38.88889
3 3 29.27350 47.00855
4 4 37.17949 57.05128
5 5 41.66667 62.39316
6 6 48.07692 66.66667
7 7 54.91453 72.00855
8 8 58.33333 77.99145
9 9 64.10256 81.41026
Result IC50 (mg/ml) 6.441273 3.451255
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FIGURE 2: DPPH scavenging activity of cucumber gel
SWELLING INDEX
From the results obtained by the study of swelling index it can be concluded that the gel has
good swelling capacity. The figure 3 represents the graph showing the swelling index.
FIGURE 3: Swelling index of cucumber gel
OTHER PARAMETERS
pH- The pH of the formulation was found to be 6.4 which was suitable to use on skin as it
comes within the range of skin pH.
Spreadability The gel formulation spreads smoothly on a clean even glass plate with
minimum pressure without the presence of any solid or gritty particles.
Homogenicity The prepared gel formulation was homogenous as on visual inspection no
aggregates were observed in the formulation.
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CONCLUSION
From the above study it can be concluded that the formulated cucumber gel is effective in
protecting skin against harmful UV radiations to some extent. The gel has the potential to
prevent against both the UVB and UVA rays as it has been observed that it has significant
SPF value and UVA/UVB ratio respectively. As the SPF obtained with a single natural
ingredient is not sufficient enough to prevent the body from harmful UV rays that is why
mostly all the commercial herbal sunscreen preparations include physical sun protecting
agents (such as titanium dioxide or zinc oxide etc.) in order to provide extra protection. The
formulation was also found to possess significant antioxidant activity which may contribute
to its sunscreen activity. In addition to the above mentioned parameters the sunscreen gel was
found to have good swelling capacity and homogenecity along with good spreadability and
pH in the range of skin. Thus the cucumber extract can be used topically not only as
moisturizing and skin rejuvenating agent but also has sunscreen effect.
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